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Fathead minnow VTG gene and protein expression after EE2 exposure

Published by U.S. EPA Office of Research and Development (ORD) | U.S. Environmental Protection Agency | Metadata Last Checked: August 02, 2025 | Last Modified: 2021-01-07
For decades, induction of vitellogenin (VTG), a yolk precursor protein, in male fish has been utilized as an indicator of environmental exposure to estrogenic compounds. VTG has been measured by a variety of enzyme-linked immunosorbent assays (ELISAs). Recently, an ELISA targeting VTG from a variety of environmentally or regulatory relevant fish species was developed which detects VTG in serum, plasma, whole tissue homogenate and outer mucus coat (TECO Medical Group, Switzerland). Detection of VTG in mucus presents a non-invasive method of monitoring endocrine disruption. As the scientific community strives to reduce animal testing, this method could provide a reliable, minimally invasive sample for routine monitoring studies. The objective of this study was to characterize VTG protein in male fathead minnow (Pimephales promelas) mucus compared with more conventional measures in plasma and mRNA isolated from liver. VTG expression changes have frequently been studied in fathead minnows exposed to the well-characterized endocrine disrupting compound, 17α-ethinylestradiol (EE2). To assess the intensity and duration of changes in mucus VTG concentrations, fathead minnows were exposed to EE2 for 7 days with a subsequent depuration period of 14 days. The exposure was conducted in a flow-through system to maintain a consistent concentration of EE2 at a nominal EC50 concentration of 2.5 ng/L and high concentration of 10 ng/L as a positive control. Mucus, plasma and liver were sampled at regular intervals throughout the study. Relative abundance of vtg mRNA increased after 2 days of exposure and disappeared after 4 days of depuration. Vitellogenin protein expression increased after 2 days of exposure and remained elevated throughout the depuration period. The pattern of protein expression in mucus was the same as plasma indicating mucus can be used as a non-invasive sample for measuring VTG in fish. Sampling mucus allows researchers to collect more samples in less time during one sampling event without sacrificing animals. Additionally, the same population may be repeatedly sampled, which is potentially useful to monitor remediation efforts or for endangered species. This dataset is associated with the following publication: See, M.J., D. Bencic, R. Flick, J. Lazorchak, and A. Biales. Characterization of vitellogenin concentration in male fathead minnow mucus compared to plasma, and liver mRNA. AQUATIC TOXICOLOGY. Elsevier Science Ltd, New York, NY, USA, 236: 113428, (2022).

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